RNA interference (RNAi) is now an umbrella term referring to post-transcriptional gene silencing mediated by either degradation or translation arrest of target RNA. This process is initiated by double-stranded RNA with sequence homology driving specificity.
RNA interference (RNAi) is an evolutionarily conserved post-transcriptional gene silencing (PTGS) mechanism mediated by double-stranded RNA (dsRNA). The dsRNA is processed into small duplex RNA molecules of approximately 21-22 nucleotides (nts) termed small interfering RNAs (siRNAs) by a RNase III enzyme called Dicer. Interaction of siRNAs with a multi-protein complex, termed the RNA-induced silencing complex (RISC), results in sequence specific association of the activated RISC complex with the cognate RNA transcript. This interaction leads to sequence-specific cleavage of the target transcript.
Originally discovered in Caenorhabditis elegans, the study of RNAi in mammalian cells has blossomed in the last couple of years with the discovery that introduction of siRNA molecules directly into somatic mammalian cells circumvents the non-specific response vertebrate cells have against larger dsRNA molecules. Emerging as a powerful tool for reverse genetic analysis, RNAi is rapidly being applied to study the function of many genes associated with human disease, in particular those associated with oncogenesis and infectious disease. Use of siRNA as a tool is advancing in almost every field of biomedical research, but some of the most dynamic and exciting applications of siRNA are in cancer research.
Almost all human cancers have accumulated multiple genetic lesions including oncogenes. It is often unknown whether an oncogene is continuously required for tumorigenesis. Furthermore, it is very difficult to target an essential oncogene with drugs without affecting the corresponding nonmutated protooncogene or related factors. RNA interference and the application of small interfering RNAs in mammalian cell culture provide new tools to examine the role of oncogenes in tumor development.
The Applicant has recently cloned a testis specific gene SPAG9 localized on human chromosome 17. It contains coiled coil domains and a leucine zipper motif encoding a protein consisting of 766 amino acids; and has been assigned to UniGene cluster Hs. 129872. Functional analysis of SPAG9 revealed that SPAG9 may have role in one or more events leading to fertilization. Southern hybridization studies suggested that human genome contains single copy of SPAG9 gene having 19 exons. The exons sequence length of SPAG9 varies from 39 to 333. The Applicant sequenced SPAG9 (CAA62987) gene the same bears SEQ ID 17 which encodes the polypeptide (766 aa) and the same bears SEQ ID 18.
Further, based on the above and upon further investigations found that the SPAG9 mRNA is expressed exclusively in normal testis tissue whereas SPAG9 is expressed in a majority of tumors (cancer) and transformed cell lines namely: testis, kidney, uterus, nervous tissue, eye, pituitary, colon, skin, lung, placenta, stomach, urinary bladder, leukopheresis, breast, vulva, pharynx, placenta, bone, prostate and liver.
There is increasing evidence for an immune response to cancer in humans, as demonstrated in part by the identification of autoantibodies against a number of intracellular and surface antigens detectable in sera from patients with different cancer types. The generation of antibodies against SPAG9 in tissues other than testis made the applicant investigate this aspect further and now, the Applicant has now developed novel sequences that are capable of targeting SPAG9 in cancerous tissues.